The long-term goals of this project are to elucidate the function and regulation of protein phosphorylation in the control of cell growth, differentiation, survival, and metabolism in response to various growth factors, such as EGF and insulin. Phosphatidylinositol-3 kinase (PI3K) is a critical component in the growth factor signaling pathway and plays an important role in cell growth and survival. A key downstream effector of PI3K is the AKT/PKB protein kinase. AKT activation requires the phosphorylation of two key residues, T308 by PDK1 and S473 by PDK2. The identity of PDK2 has remained elusive until the recent identification of TORC2 (TOR complex 2) as a candidate PDK2. Recent studies have established that TORC2 plays a key role in the phosphorylation of AKT S473 and is stimulated by the PI3K pathway. Dysregulation of the PI3K pathway is associated with many human diseases, such as cancers and hypertrophic disorders. We have identified Sin1 as a novel TORC2 component required for complex formation and kinase activity. The major goal of this proposal is to investigate the molecular mechanism of TORC2 regulation by insulin and PI3K. The specific aims of the proposal are: 1. To determine the functional significance of Sin1 phosphorylation in regulation of TORC2 2. To determine the regulation of Sin1 protein levels by Rictor and mTOR 3. To investigate the mechanisms of TORC2 activation by insulin and PI3K